The binding of erythropoietin to a specific receptor on the surface of erythroid progenitors stimulates erythropoiesis and results in a number of molecular events including the increased production of GATA- 1, an erythroid specific transcription factor, and activation of globin genes. We initiated studies on the erythropoietin receptor in order to probe early events in erythropoiesis. Analysis of transcription of the erythropoietin receptor revealed much of the transcription activity is contained within the proximal 150 bp 5' which contains binding sites for GATA-1 and SP1 transcription factors that are necessary for promoter activity. Protein-DNA binding studies suggest possible interactions between GATA-1 and SP1. Increasing the distance between their binding sites by half turns of the DNA helix (+5, +10, +15 and +21 bp) revealed phasing in transcription activation as measured in transient transfection studies, consistent with this hypothesis. We are now studying the interactions among these transcription factors. In transgenic mice containing a 15 kilobase human erythropoietin receptor gene, transgene expression was detected in the brain, both in embryos and adults, suggesting that the erythropoietin receptor might function in the brain. Examination of early embryonic brain (day 10) showed transcripts of the endogenous erythropoietin receptor at significant levels which decreased by several logs as the embryo matured (day 16) and largely disappeared after birth. Both the endogenous and transgene were expressed in embryonic and adult hematopoietic tissue. We have now recapitulated mouse erythropoietin receptor gene expression using a 77 kilobase human erythropoietin receptor gene construct with appropriate regulation of brain expression suggesting that embryonic brain expression may also occur during human development. Using an erythropoietin receptor promoter/beta-galactosidase reporter transgene suggests possible localization in the neurotube at day 9 and neurotube mRNA also contained endogenous erythropoietin receptor transcripts. We have isolated several erythropoietin receptor clones from a human brain cDNA library that suggest the erythropoietin receptor mRNA in the brain may exist as an alternate form. We have identified two human neuronal cell lines which express the erythropoietin receptor and should be useful in determining its possible functional importance in proliferation and differentiation. These studies suggest that the erythropoietin receptor expression is not restricted to hematopoietic cells and may also play a role in the development of select non- hematopoietic tissues.